Tweet
Does anyone sequence DNA samples that have gone through siRNA, shRNA, or miRNA screening? If yes, can someone explain how it is done on the Illumina?
Basically what I know is a typical protocol will have the siRNA/shRNA/miRNA insert in a vector or virus particle which is transfected into cells, cells go through a screening and what is left in the end are viable cells with the siRNA/shRNA/miRNA integrated into the genome. Then one wants to make sure the target gene has been silenced, so they sequence the DNA from these cells to confirm if the siRNA/shRNA/miRNA has been stably integrated into the genome and to find that it is a true match to the target gene for silencing.
But how is the sequencing aspect done on the Illumina sequencer? I have not found a good guide. Is it single-end sequencing or paired-end?
Basically what I know is a typical protocol will have the siRNA/shRNA/miRNA insert in a vector or virus particle which is transfected into cells, cells go through a screening and what is left in the end are viable cells with the siRNA/shRNA/miRNA integrated into the genome. Then one wants to make sure the target gene has been silenced, so they sequence the DNA from these cells to confirm if the siRNA/shRNA/miRNA has been stably integrated into the genome and to find that it is a true match to the target gene for silencing.
But how is the sequencing aspect done on the Illumina sequencer? I have not found a good guide. Is it single-end sequencing or paired-end?
Comment